Hypertension is generally defined as the elevation of the systolic and/or diastolic arterial blood pressure above a nominal value of 140/90 mm Hg. Diseases associated with hypertension include arteriosclerosis, hypertensive renal failure, stroke, congestive heart failure and myocardial infarction. Although numerous methods of treatment have been found to be effective in the reduction of arterial blood pressure, the etiology of essential hypertension remains essentially unknown. A genetic predisposition to hypertension is generally accepted, but the number of different drugs which have been found effective in the treatment of hypertension, and the fact that these drugs seem to operate by eliciting different pharmacological responses, suggests that there may be different primary causes for essential hypertension.
A number of studies have suggested that one or more circulating factors may play a role in the genesis or the maintenance of hypertension [See: Wright et al., A Hypertensive Substance Found in the Blood of Spontaneously Hypertensive Rats; Life Sci. 1984; 34:1521-1528; Dahl et al., Humoral transmission of Hypertension: Evidence from Parabiosis; Circ. Res. 1969; 24/25 (Supp. I): 21-23; Greenberg et al., Evidence for Circulating Factors as a Cause of Venous Hypertrophy in Spontaneously Hypertensive Rats; Am. J. Physiol. 1981; 241:H421-H430; Tobian et al., A Circulating Humoral Pressor Agent in Dahl S Rats with Salt Hypertension; Clin. Sci. 1979; 57:345s-347s; Zidek et al., Humoral Factors in the Pathogenesis of Primary Hypertension: Klin. Wochenschr. 1985; 63 (Supp. II) D:94-96; Hirata et al., Hypertension Producing Factor in the Serum of Hypertensive Dahl Salt-Sensitive Rats; Hypertension 1984; 6:709-716]. For example, in parabiosis and cross-circulation experiments, an increase in blood pressure could be induced in normotensive animals by exposure to blood from hypertensive animals. The subcutaneous injection of erythrocyte-associated factor obtained from spontaneously hypertensive rates (SHR) has been shown to induce hypertension in normotensive Wistar-Kyoto (WKY) rats and an increase in blood pressure can be induced in normotensive, salt insensitive Dahl rats by injection of serum from hypertensive, salt-sensitive Dahl rats.
There have also been reports of circulating factors in both hypertensive rats and hypertensive humans which increase intracellular calcium. [See: Banos et al., Two Factors Associated with Increased Uptake of Calcium in Platelets from Essential Hypertensive Patients; Clin. Exp. Hypertens. 1987; 9:1515-153; Zidek et al., Effect of Plasma from Hypertensive Subjects on Ca Transport in Permeabilized Human Neutrophils; Clin. Sci. 1988; 74:53-56; Linder et al., Effects of a Circulating Factor in Patients with Essential Hypertension on intracellular Free Calcium in Normal Platelets; N. Eng. J. Med. 1987; 316:509-513; Bruschi et al., Cytoplasmic Free Ca is Increased in the Platelets of Spontaneously Hypertensive Rats and Essential Hypertensive Patients; Clin. Sci. 1985; 68:179-184; Wright et al., Stimulation of Aortic Tissue Calcium Uptake by an Extract of Spontaneously Hypertensive Rat erythrocytes Possessing Hypertensive Properties; Can. J. Physiol. Pharmacol. 1986; 64:1515-1520]. Since vascular tone is influenced by the level of intracellular calcium, factors which increase blood pressure and factors which increase intracellular calcium may be related. There has been accumulating evidence suggesting the involvement of calcium regulating hormones in some forms of hypertension [See: L. M. Resnick, Am. J. Med. 82 (Supp. 1B), 16 (1987)]. Parathyroid hormone (PTH) is a calcium regulating hormone. Thirty percent or more of essential hypertensive patients fall into a subgroup characterized by increased levels of immunoreactive parathyroid hormone (ir-PTH). [See: Laragh et al., Kidney Int. 34, (Supp. 35), S162 (1988)]. An increase in PTH levels has been reported in SHR rats [See: McCarron et al., Hypertension 3 (Supp. 1), I162 (1981)] and it has been observed that hyperparathyroid patients often exhibit hypertension, the severity of which can, in most cases, be reduced by parathyroidectomy [See: Hellstrom et al., Brit. J. Urol. 30, 13 (1958)]. Similar results from parathyroidectomy have also been reported in SHR rats. [See: Schleiffer et al., Jap. Circ. J. 45, 1272 (1981)]. Various investigators have suggested that PTH contributes to the development of essential hypertension, although exogenous administration of PTH causes a reduction in blood pressure in mammals and other vertebrates [See: Pang et al., Gen. Comp. Endocrinol. 41, 135 (1980)]. The vasodilating action of PTH is also related to a specific region of the molecule separate from the region mediating hypercalcemic effects [See: Pang et al., Endocrinology, 112, 284 (1983)]. PTH has also been shown to inhibit calcium entry into vascular smooth muscle [See: Pang et al., Life Sci., 42, 1395 (1988)] through L-type calcium channels [Wang et al. FEBS, Vol. 282, No. 2, pp. 331-334 (1991)]. This paradox is further heightened by the fact that hypertensive patients with increased PTH levels exhibit decreased serum ionized calcium levels [See: Resnick et al., New Engl. J. Med., 309, 888 (1983); Hvarfner et al., Acta Med. Scand. 219, 461 (1986)]. It would be expected that the serum ionized calcium levels would be elevated if PTH were primarily elevated.
The existence of a circulating factor in the blood of the SHR rat was confirmed by the studies reported in Am. J. Hypertens., 2, 26-31 (1989). In these studies, an increase in the blood pressure of WKY and SD rats when plasma from SHR rats was injected into the normotensive rats either by infusion or by bolus injection was shown. In addition, it has been shown that the uptake of 45Ca by sections of the tail artery of a rat, in vitro, increased in a dose-dependent manner as the concentration of SHR plasma was increased in a buffer-based medium. The results of these experiments clearly show that an increase in blood pressure and an increase in calcium uptake in the cells were both dose-dependent on the amount of SHR plasma present and available in the system. Curiously, the onset of both events was delayed, and gradual, whereas known endogenous pressor agents such as norepinephrine, angiotensin II and vasopressin have been observed to increase blood pressure quite rapidly after administration. The known endogenous pressor agents exhibit about a 1-2 minute onset in the increase of blood pressure and increase in calcium uptake in the cells whereas parathyroid hypertensive factor has a 20-30 minute delay before such onset. Another result observed in these studies was that when the infusion of SHR plasma was stopped and substituted with plasma from normotensive rats, the observed blood pressure decreased quite rapidly to the baseline. The decrease observed precluded a simple volume effect. In a related experiment, dialyzed plasma from hypertensive human subjects was infused into normotensive SD rats and shown to produce hypertension. Plasma from these subjects also increased calcium uptake in rat tail arteries in vitro. Dialyzed plasma from normotensive patients produced no significant increase in blood pressure.
The origin of the circulating factor was unknown, but the anecdotal reports that PTH was elevated in hypertensive rats suggested the parathyroid gland as a target of investigation. Parathyroidectomies of SHR rats were found to reduce blood pressure and plasma from the SHR rats which had been parathyroidectomized did not cause elevation of blood pressure in normotensive rats. Conversely, transplantation of parathyroid glands from SHR rats to normotensive Sprague-Dawley (SD) rats resulted in an increase in blood pressure and the appearance of the factor in the plasma, as shown by infusion of the isolated plasma into other normotensive rats. [Pang and Lewanczuk, Amer. J. Hypertens. 2, 898 (1989)].
On the basis of these studies, the parathyroid was determined to be the origin of the circulating factor and the name “Parathyroid Hypertensive Factor” or PHF was proposed for the substance which causes an elevation in blood pressure.
The isolation and purification of a circulating factor, having its origin in the parathyroid gland, has been demonstrated in SHR rats and in many humans having essential hypertension and is the subject matter of related patent application Ser. No. 603,745 filed Nov. 21, 1990, which is a continuation-in-part of patent application Ser. No. 327,450, filed Mar. 22, 1989, now abandoned. The disclosure of the related patent applications are incorporated herein by reference for their teachings, including the teachings of purification of parathyroid hypertensive factor.
As described in the aforementioned related patent applications, PHF has been shown to regulate extracellular calcium uptake, and can be inhibited by increases in dietary calcium levels. PHF has been isolated and a method for screening for PHF using antibodies raised against PHF have been described. PHF has a molecular weight of approximately 2,700 daltons and has the property of delayed onset of an increase in blood pressure of a normotensive rat when administered thereto, the increase in blood pressure temporally correlating with an increase in extracellular calcium uptake by vascular smooth muscle. From bioassay data, the factor in humans and rats has been found to be substantially similar.
Vascular hypertrophy has been implicated in the pathophysiology of a number of cardiovascular diseases including essential hypertension. Vascular smooth muscle proliferation could account for vascular hypertrophy and increased vascular tone. It was reported that PHF increased vascular smooth muscle cell proliferation through a mechanism independent of intracellular calcium regulation (Shan et al., Abstract in 17th Scientific Meetings of the International Society of Hypertension, Amsterdam, 7-11 Jun. 1998).
Antagonists of PHF have been found by the present inventors. The present inventors have unexpectedly found that shark cartilage, known in the art to contain a substance which inhibits tumor angiogenesis [Lee et al., Science, vol. 221, pp.1185-1187, (1983)] and to contain an anti-inflammatory component [Schinitsky U.S. Pat. No. 4,473,551], acts as an antagonist of PHF resulting in a decrease in blood pressure and affecting intracellular calcium regulation. The present inventors have also found that shark cartilage extract inhibited VSMC proliferation in SHR rats or in WKY rats induced by PHF. In view of this, shark cartilage extract according to the present invention is expected to be useful for treating hypertension and other diseases related to intracellular calcium elevation.